Regulatory aspects of glutamate metabolism in Aspergillus

The object of this study has been to extend our present knowledge of the control of gene action in eukaryotic cells. For the purpose of achieving this goal systems responsible for L-glutamate metabolism in the simple fungus Aspergillus nidulans have been chosen for investigation. These systems are L-glutamate transport, NADP L-glutamate dehydrogenase and NAD L-glutamate dehydrogenase. A mutant, unable to grow on L-glutamate and L-aspartate as a carbon or nitrogen source and lacking L-glutamate and L-aspartate transport has been isolated. This asuAl grows normally on other amino acids as sole carbon or nitrogen sources and appears to have normal uptake of all other nitrogen sources tested indicating that there is a specific uptake system for L-glutamate and L-aspartate. This system is regulated by ammonium as are a number of other unrelated systems, e.g. nitrate reductase, acetaraidase, etc. It is not clear whether this ammonium control is at the level of transcription or translation but for convenience referred to as ammonium repression. The synthesis of L-glutamate from ammonium and L-oxoglutarate is catalysed by HADP L-glutamate dehydrogenase. Mutants; unable to grow normally on ammonium and lacking NADP L-glutamate dehydrogenase activity, have been isolated. The mutants, designated gdhAl - A9, require a supplement of amino acid for normal wild type growth. In addition to losing their catalytic activity, gdhA mutants are insensitive to ammonium control of ammonium repressible systems, including glutamate transport. A model for these results has been proposed. This suggests that NADP L-glutamate dehydrogenase plays a role in ammonium repression in addition to its catalytic function. Aspergillus can utilise L-glutamate as a sole carbon source. Mutants, designated gdhBl - unable to grow on glutamate as a sole carbon source and lacking NAD L-glutamate dehydrogenase activity, have been isolated. The gdhB mutants can still grow on ammonium or L-glutamate or other amino acids as sole nitrogen sources with glucose as the carbon source. HAD L-glutamate dehydrogenase activity is under repression by carbon metabolites, with glucose but not acetate, as a source of these metabolites. It may also be inducible but this is unclear at present. If induction plays a part in the control mechanism then it is probably subordinate to repression. A mutant which grows better than the wild type with glutamate as a nitrogen source has been isolated. This gdhCl mutant is partly insensitive to glucose repression of HAD L-glutamate dehydrogenase. The gdhCl mutation is semi-dominant in the heterozygous diploid. Again a model has been proposed. This suggests that the gdhC locus codes for a regulatory product which has a positive effect on NAD L-glutamate dehydrogenase synthesis. The product of the gdhC locus is involved in repression by glucose or metabolites derived from glucose of NAD L-glutamate dehydrogenase

Genetic characterization of morphologically variant strains of Paracoccidioides brasiliensis

Molecular characterization of Paracoccidioides brasiliensis variant strains that had been preserved under mineral oil for decades was carried out by random amplified polymorphic DNA analysis (RAPD). On P. brasiliensis variants in the transitional phase and strains with typical morphology, RAPD produced reproducible polymorphic amplification products that differentiated them. A dendrogram based on the generated RAPD patterns placed the 14 P. brasiliensis strains into five groups with similarity coefficients of 72%. A high correlation between the genotypic and phenotypic characteristics of the strains was observed. A 750 bp-RAPD fragment found only in the wild-type phenotype strains was cloned and sequenced. Genetic similarity analysis using BLASTx suggested that this RAPD marker represents a putative domain of a hypothetical flavin-binding monooxygenase (FMO)-like protein of Neurospora crassa.FiocruzBritish Council Progra